Synthesis, spectroscopic characterization, stability assessment and DNA-binding of new 2,6-piperidinedione derivatives.

This work reports on structural characterization of new antineoplaston (ANP) representatives, namely 3-(benzoylamino)-2,6-piperidinedione (BPD), 3-(4-methoxybenzoylamino)-2,6-piperidinedione (MPD) and 3-(p-nitrobenzoylamino)-2,6-piperidinedione (NPD). These compounds were prepared by reacting N-(4-substituted benzoyl)-glutamines with N-hydroxysuccinimide to afford the corresponding esters, which were heated to produce the corresponding 2,6-piperidinedione (PD) compounds. Non-destructive analytical procedures such as 1H NMR and NIR analyses confirmed the postulated chemical structures of these PD compounds. HPLC chromatograms at an ambient temperature or from solutions preheated at 30, 40 or 60 degrees C displayed only a single peak for each compound. Combination of heat with pH modification had virtually no effect on the obtained peaks, thus attesting to the stability and purity of these compounds. MS analysis displayed molecular mass ions indicative of BPD, MPD and NPD at m/z 233.4, 263.2 and 278.3, respectively. The fragmentation patterns using MS/MS analyses conformed to the structural and molecular formulae of the prepared compounds. Furthermore, preliminary biological assessments showed the capacity of these compounds to bind to the DNA. NPD, but not BMP or MPD, had a superior affinity to the DNA than the prototype ANP-A10.

Immune modulatory potentials of antineoplaston A-10 in breast cancer patients.

Antineoplastons are naturally occurring cytodifferentiating agents. Chemically, they are medium and small sized peptides, amino acid derivatives and organic acids, which exist in blood, tissues and urine. Antineoplaston A-10 (3-phenylacetylamino-2,6-piperidinedione) is the first chemically identified antineoplaston. Previously we have shown a strong inverse association of urinary antineoplaston A-10 with breast cancer. This study is designed to evaluate neutrophil apoptosis in patients with breast cancer at time of diagnosis and to correlate urinary antineoplaston A-10 levels with neutrophil apoptosis and to describe the direct effect of A-10 in vitro on neutrophil apoptosis in breast cancer patients. The participants were patients with a histologically confirmed diagnosis of breast cancer. Only those cases without previous treatment for breast cancer were included. Neutrophil apoptosis was assessed in breast cancer patients both morphologically and by DNA fragmentation and studied relative to healthy controls. Antineoplaston A-10 was measured using high performance liquid chromatography in urine samples collected from the patients. Urine samples from normal women served as controls. Direct effect of antineoplaston A-10 on neutrophil apoptosis was tested in vitro after adding A-10 at a concentration of 10 ng/ml to the cellular suspensions of breast cancer patients. Non-treated samples served as controls. Significantly higher neutrophil apoptosis levels were detected among patients with breast cancer with a P value <0.001. Urinary antineoplaston A-10 level is significantly negatively correlated with high apoptosis levels (P<0.0001). In vitro, antineoplaston A-10 was found to inhibit significantly the neutrophil apoptosis with a P value <0.0001. These findings confirm the presence of immune defects among patients with breast cancer and such results should stimulate the development of new strategies to induce and augment immunity for the treatment of breast cancer. Antineoplaston A-10 may provide rational basis for designing trials to employ its immune modulatory potentials as adjuvant therapy in breast cancer patients.

Potential utility of antineoplaston A-10 levels in breast cancer.

Antineoplastons, first described by Burzynski, are naturally occurring peptides and amino acid derivatives, which control neoplastic growth. Antineoplaston A-10 (3-phenylacetyl amino-2, 6-pepridinedione) is the first chemically identified antineoplaston. Here we describe the potential utility of antineoplaston A-10 as a predictive test for breast cancer. Antineoplaston A-10 level was measured in the urine of 31 breast cancer patients and 17 normal women using high performance thin layer chromatography (HPTLC). Significantly lower antineoplaston A-10 levels were detected among patients with breast cancer with a P value <0.001. These data suggest a strong inverse association of urinary antineoplaston A-10 level with breast cancer. Such finding was the stimulus for further investigations of antineoplaston A-10 levels in some benign as well as other malignant diseases to determine the utility of this approach as a predictive test for women who are at risk of developing breast cancer.

Novel piperidinedione analogs as inhibitors of breast cancer cell growth.

We previously reported the utility of antineoplaston-A10 (3-phenylacetylamino-2,6-piperidinedione) as an endogenous cancer protector and immune modulator in breast cancer patients (Cancer Lett., 2000, 157, 57). In this study, four new piperidinedione A10 analogs were synthesized and tested for their antimitotic activity on a human breast cancer cell line against the prototype A10 and the antibreast cancer drug tamoxifen. Moreover, the DNA binding capacity of such compounds was evaluated against A10, (E)-3-(4-Nitrocinnamoylamino)-2,6-piperidinedione "3B" and (E)-3-(4-hydroxycinnamoylamino)-2,6-piperidinedione "3D" were several-fold more potent antiproliferative agents than A10 and tamoxifen. They also had significantly higher capacity to bind DNA than A10. Conversely, (E)-3-(cinnamoylamino)-2,6-piperidinedione "3A" and (E)-3-(4-methoxycinnamoylamino)-2,6-piperidinedione) "3C" had weaker biological profiles than the lead compound A10. Detailed synthetic, spectroscopic, and biological data are reported.